Blood transfusion is a life-saving intervention, yet the treatment is not without complications. One event that can occur is generation of an immune response directed toward the blood donor's genetic background (alloimmunization). This phenomenon can lead to adverse consequences, such as refractoriness to platelet transfusion, a particular problem for patients requiring multiple transfusions for support, such as oncology patients after myeloablative therapy. In addition, allosensitization of patients by transfusion, fo example dialysis patients, can complicate subsequent attempts to find a suitable organ for transplantation due to difficulties with HLA cross-matching. A recent genome-wide association study (GWAS) identified a single nucleotide polymorphism (SNP) within the NXPH2 gene that was associated with strong protection from alloimmunization after pregnancy (odds ratio 0.51). The minor allele variant frequency was 16.7% in a population of Caucasian women, meaning that approximately 30% of the women studied carried at least one copy of the protective variant. This proposal will develop the necessary reagents and perform the preliminary studies needed to confirm and extend the association between NXPH2 and alloimmunization and determine if NXPH2 plays a functional role in immune modulation, opening a new field of research. We will develop PCR-based assays to measure the presence of four SNPs associated with protection from alloimmunization, and these new assays will be used to test a replicate cohort of subjects who were or were not alloimmunized after blood transfusion. This will determine whether or not the findings of the GWAS can be replicated in an independent cohort. Additionally, the level of NXPH2 protein in subjects with and without alloimmunization after exposure (via pregnancy or transfusion) will be measured, with the hypothesis that higher levels of NXPH2 will protect from alloimmunization. Finally, functional studies will be performed to determine if NXPH2 directly interacts with immune cells and if so with which cells the protein interacts. This proposal will open a new field of research if the findings of the original GWAS can be replicated. NXPH2 has no known role in the immune system, and the ability to manipulate this pathway could prove useful in up- or down-regulating immune responses. This exploratory grant will determine if NXPH2 interacts with the immune system and will develop reagents for further study of these interactions.